1. Technical Field
The present invention relates generally to fusion proteins made from a synthetic chimeric gene construct comprising sequences from K26, K39, and K9 and their use in the diagnosis and treatment of leishmaniasis.
2. Description of the Related Art
Leishmania organisms are intracellular protozoan parasites of macrophages that cause a wide range of clinical diseases in humans and domestic animals, primarily dogs. In some infections, the parasite may lie dormant for many years. In other cases, the host may develop one of a variety of forms of leishmaniasis. For example, the disease may be asymptomatic or may be manifested as sub-clinical visceral leishmaniasis, which is characterized by mild symptoms of malaise, diarrhea and intermittent hepatomegaly. Patients with sub-clinical or asymptomatic disease usually have low antibody titers, making the disease difficult to detect with standard techniques. Alternatively, leishmaniasis may be manifested as a cutaneous disease, which is a severe medical problem but is generally self-limiting, or as a highly destructive mucosal disease, which is not self-limiting. Finally, and most seriously, the disease may be manifested as an acute visceral infection involving the spleen, liver and lymph nodes, which, if left untreated, is generally a fatal disease. Symptoms of acute visceral leishmaniasis include hepatosplenomegaly, fever, leukopenia, anemia and hypergamma-globulinemia.
Three main clinical variants of this disease are known: cutaneous, mucocutaneous, and visceral. Cutaneous leishmaniasis can manifest itself as a single skin ulceration at the site of the sandfly bite appearing soon after infection or months later as disseminated lesions. Mucocutaneous syndrome develops as the cutaneous form, but progresses months or years later to lesions of the mouth, nose, or pharynx. The major long-term effects of cutaneous and mucocutaneous disease are scarring. Visceral leishmaniasis has an incubation period of 3-6 months and involves the reticuloendothelial system.
Clinical manifestations of visceral leishmaniasis include enlargement of the liver and spleen, fever, anemia, and weight loss. In the absence of treatment, symptomatic visceral disease often ends in death. In recent years, the coexistence of HIV and Leishmania species causing visceral disease has resulted in several hundreds of cases of dually infected individuals (Berman, J. D., (1997) Clin. Infect. Dis. 24:684). The World Health Organization recently estimated in 2000 that leishmaniasis affects people in 88 countries, with 350 million at risk of contracting the disease and about two million new cases each year. The devastating impact of this disease is exemplified by the recent epidemic of visceral leishmaniasis in the Sudan, which claimed an estimated 100,000 lives (Seaman, J., et al. (1996) Int. J. Epidemiol. 25:862). This disease is frequently a threat in military operations, as demonstrated by the outbreak of viscerotropic leishmaniasis during the Gulf War (Magill, J., et al. (1993) N Engl J Med 328:1383).
Leishmaniasis is a serious problem in much of the world, including Brazil, China, East Africa, India and areas of the Middle East. The disease is also endemic in the Mediterranean region, including southern France, Italy, Greece, Spain, Portugal and North Africa. The number of cases of leishmaniasis has increased dramatically in the last 20 years, and millions of cases of this disease now exist worldwide. About 2 million new cases are diagnosed each year, 25% of which are visceral leishmaniasis. There are, however, no vaccines or effective treatments currently available.
Leishmaniasis is caused by several species of Leishmania. These unicellular organisms of the order Kinetoplastida are related to trypanosomes, the causative organisms of Sleeping Sickness in Africa and Chagas' disease in South America. Leishmania parasites commonly exist in two distinct forms, the motile promastigote of the insect vector and the sessile amastigote present in the mammalian host. Promastigotes are transmitted to humans by the bite of infected phlebotomine sandflies, which are found throughout the world's inter-tropical and temperate regions. Upon delivery into the mammalian host, promastigotes infect macrophages of the reticuloendothelial system and transform into amastigotes.
Accurate diagnosis of leishmaniasis is frequently difficult to achieve. There are 20 species of Leishmania that infect humans, including L. donovani, L. chagasi, L. infantum, L. major, L. amazonensis, L. braziliensis, L. panamensis, L. mexicana, L. tropica, and L. guyanensis, and there are no distinctive signs or symptoms that unambiguously indicate the presence of Leishmania infection.
Parasite detection methods have been used, but such methods are neither sensitive nor clinically practical. Current skin tests typically use whole or lysed parasites. Such tests are generally insensitive, irreproducible and prone to cross-reaction with a variety of other diseases. In addition, the preparations employed in such tests are often unstable.
Thus, there is a need for improved methods for the detection of Leishmania infection. For example, there is a need in the art for more sensitive and specific methods for detecting Leishmania infection, and for identifying those asymptomatic Leishmania infections that are likely to progress to acute visceral infections. The present invention fulfills these needs and further provides other related advantages.